Despite an increasingly negative attitude towards smoking and intensified public campaigns and legislation against smoking, virtually no further reduction in smoking has occurred in this country during the 1990's. Based on the 1996 National Household Survey on drug abuse, an estimated 68.8 million Americans used tobacco products. Therefore, tobacco represents one of the most widely abused substances. Nicotine is believed to be the primary addictive constituent in tobacco. Both epidemiological and molecular studies have implied that many genes respond to nicotine stimulation and there exists significant genetic differences between males and females in response to nicotine administration. Even though numerous studies have been conducted to investigate how a gene of interest is modulated by nicotine using both in vivo and in vitro systems, only a limited number of systematic studies were reported on gene expression profile during chronic exposure to nicotine. We hypothesize that distinct genes and pathways could be identified by a genome-wide functional genomic approach, such as microarray technique. Specially, the aims of this proposal are: 1) to identify novel genes associated with nicotine dependence and nicotine withdrawal in male and female adult rats using a high-density cDNA microarray generated and annotated in the laboratory; 2) to characterize several most promising genes identified from Aim 1 in detail at DNA, RNA, and protein levels with various conventional molecular techniques using both in vitro and in vivo systems; 3) to identify novel signaling pathways involved in nicotine dependence and nicotine withdrawal in rats using the pathway- focused microarray developed in the laboratory; and 4) to implement and develop databases and statistical methodologies for managing and analyzing microarray data generated from Aims 1-3. Such a global perspective may provide new direction to develop preventive and treatment strategies to nicotine dependence and nicotine withdrawal in human smokers. The completion of the proposed studies will advance our understanding of complex gene expression patterns associated with nicotine dependence and nicotine withdrawal in rats and in human postmortem brain tissues.